Ordered recruitment of histone acetyltransferases and the TRAP/Mediator complex to thyroid hormone-responsive promoters in vivo

Abstract

Transcriptional coactivators implicated in gene activation by the thyroid hormone receptor (TR) include members of the p160͞ steroid receptor coactivator (SRC) family of proteins, p300, and the multisubunit TR-associated protein (TRAP)͞Mediator complex. We investigated the temporal recruitment of these cofactors to mammalian thyroid hormone (T3)-responsive promoters in vivo. We show that upon T3 treatment, TR recruits all three types of coactivators to specific promoters in at least two sequential steps: p160͞SRC proteins and p300 are recruited first and rapidly induce histone acetylation, followed by the recruitment of the TRAP͞ Mediator complex. Interestingly, inhibition of histone deacetylase activity with trichostatin A elicited a more rapid promoter recruitment of the TRAP͞Mediator complex but not p160͞SRC proteins. T3-dependent gene expression assays indicate that all three coactivators are targeted to a promoter before significant activation occurs. These findings thus suggest that histone acetylation may be a prerequisite for TRAP͞Mediator recruitment and function at specific T3-responsive mammalian promoters.T hyroid hormone receptors (TRs) are members of the nuclear receptor (NR) superfamily and regulate transcription by recruiting specific coregulatory protein complexes to target gene promoters (reviewed in refs. 1 and 2). TR binds to target promoters as a heterodimer with the retinoid X receptor (RXR) and recruits transcriptional coactivators in the presence of thyroid hormone (T3) and corepressors in the absence of T3. Coactivators that directly interact with liganded TR and facilitate transactivation include members of the p160͞steroid receptor coactivator (SRC) family (reviewed in refs. 3 and 4). These proteins are thought to function in part by associating with potent histone acetyltransferases (HATs) such as p300͞CREB binding protein and ultimately target the HAT activity to promoter-bound TR resulting in acetylation of nucleosomes. Additionally, some p160͞SRC family members contain intrinsic HAT activity (5, 6), further supporting a functional role for these factors in chromatin modification.A second type of TR coactivator is the multisubunit TRassociated protein (TRAP)͞Mediator complex (7, 8), which appears to bind liganded TR through a single subunit termed TRAP220 (9, 10). In vitro transcription assays show that TRAP͞ Mediator significantly enhances TR-dependent transcription on nonchromatin DNA templates (7,11,12), thus suggesting that the complex functions by positively influencing the basal transcription machinery, possibly by directly facilitating the recruitment of RNA polymerase II. Consistent with this view, several TRAP͞Mediator subunits share homology with proteins in yeast Mediator, a large coactivator complex that directly associates with both transcriptional activators and the yeast RNA polymerase II holoenzyme (13). Most TRAP͞Mediator subunits also have been identified in other large transcriptional coregulatory complexes, including NAT, DRIP, ARC, and CRSP (reviewed in ref. 14)...

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